PROJECT SUMMARY (See instmctions): Sepsis is a disease process representing the systemic response to severe infection. The infectious insult activates immune cells of the host, including neutrophils, monocytes/macrophages, and lymphocytes. Early activation of immune cells triggers a pro-inflammatory response that contributes to eradication of the invading microorganism(s). If the pathogens are not eradicated, sepsis may progress to severe sepsis (sepsis plus organ dysfunction, such as lung injury and respiratory failure) and septic shock (severe sepsis plus refractory hypotension and circulatory failure), which frequently leads to death. Once the invading microorganism(s) are eradicated, then resolution of the immune response is critical, as continued systemic inflammation will lead to organ injury. Unfortunately, this early pro-inflammatory response may also be followed by a later state of immunoparalysis, due in part to apoptosis of immune effector cells. The cascade of biologic events that occur during sepsis is complex, and therapeutic strategies need to be tailored depending upon the stage of sepsis at the time of diagnosis. Multipotent mesenchymal stromal cells (MSCs) are considered to be a promising platform for cell-based therapy. MSCs are known to have immunomodulatory properties, and recently it has been suggested that MSCs are beneficial during the early proinflammatory stage of cecal ligation and puncture in mice. We hypothesize that MSCs, when administered to mice after the onset of polymicrobial or single organism sepsis, will adapt to the specific stage of sepsis and lead to an improved outcome. Prior studies in our laboratory demonstrated that heme oxygenase (H0)-1-derived carbon monoxide (CO), a known anti-inflammatory molecule, also has anti-microbial properties during sepsis. Thus, we also hypothesize that conditioning MSCs with CO ex vivo will improve their function In vivo, and increase the beneficial effects of MSCs after the onset of sepsis in mice. Gaining insight into the mechanisms responsible for this enhanced response of MSCs conditioned with CO will allow us to further understand MSCs as a cell-based therapy, and to develop novel therapeutic strategies for this devastating disease process. MSCs CO conditioning will also be studied in mice with a humanized immune system. Finally we will advance the understanding of the human immune response to sepsis by assessing phagocytic function of neutrophils and monocytes harvested from patients with sepsis or sepsis plus acute lung injury, compared with cells from control patients. We will determine whether the administration of MSCs CO ex vivo can improve phagocyte function. These functional assays will be correlated with patient outcome.